Axon regeneration is a fundamental and conserved property of neurons. A key question in the field is to discover what determines the regenerative capacity of injured neurons. The overarching goal of this proposal is to discover how three novel factors-Notch, Mint/LIN-10, and Rab6-function to inhibit regeneration. The three central hypotheses are: 1) that Mint/LIN- 10 and Rab6 work together in injured neurons to inhibit regeneration; 2) that they do so by promoting the intracellular trafficking of one or more inhibitory factors; and 3) that Notch signaling regulates one or more components of this inhibitory system. These hypotheses are supported by substantial preliminary data, and will be tested in the three Aims. 1) Define the regeneration function of Mint and Rab6 in motor and sensory neurons. This aim will establish how Mint/LIN-10 and Rab6 work together to inhibit axon regeneration, and how they function in different neurons. 2) Analyze Mint/LIN-10 binding partners in regeneration. This aim will identify the effector molecules that mediate Mint/LIN-10's regeneration function. 3) Investigate the cell biology of Mint, Rab6, and effectors in regulating axon regeneration. This aim will use in vivo real time imaging to determine the link between intracellular trafficking of these inhibitory factors and axon regeneration. These aims use a number of innovative techniques, and the proposal as a whole provides a major conceptual advance in our understanding of protein function and intracellular trafficking in relation to axon regeneration. By describing a novel neuronal mechanism that inhibits the ability of neurons to regenerate after injury, this proposal has the potential to significantly affect and improve curren efforts to find new treatments for damaged axons.